Role Of Circular Rna Asph In Macrophage Polarization And Response In Sepsis, Tan Ze Wang

Electronic Thesis and Dissertation Repository

Circular RNAs (circRNAs), a novel non-coding RNA species generated by back-splicing, has been shown to participate in gene regulation of leukocytes. Our previous RNA sequencing results show circular RNA ASPH (circASPH) to be highly expressed in peripheral blood mononuclear cells of sepsis patients at the start of intensive care. Macrophages, as ubiquitous innate immune cells, are responsible for the recruitment of other immune cells at sepsis onset. This work investigates the role of circASPH in the regulation of macrophage polarization in sepsis. Using an in vitro THP-1 cell model, it was found that circASPH levels peaked after 24 h of …

Characterizing The Function Of B Cells That Accumulate In The Inflamed Central Nervous System In Anti-Myelin Autoimmunity, Lika Chowdhury

Electronic Thesis and Dissertation Repository

While the role of autoimmune T cells has been extensively studied in anti-myelin

autoimmunity, little is known about the function of B cells in multiple sclerosis (MS), a chronic inflammatory disease of the central nervous system (CNS). B cells form clusters with T cells in the meninges directly adjacent to demyelinating lesions. Previous studies have shown that disease progression is dependent on the depletion of specific populations of B cells, but it is not clear which contributes to pathology or how. The purpose of this thesis is to characterize the population of meningeal B cells to determine how they differ …

The Role Of Reactive Oxygen Species In The Accumulation Of Driver Mutations In B Cell Acute Lymphoblastic Leukemia, Mia P. Sams

Electronic Thesis and Dissertation Repository

B cell acute lymphoblastic leukemia (B-ALL) is the most prevalent type of cancer in young children and is associated with recurrent mutations and high levels of reactive oxygen species (ROS). The antioxidant N-acetylcysteine was tested for its ability to prolong lifespan of a mouse model of B-ALL and reduce frequency of mutations. Mice treated with 1g/L of N-acetylcysteine in drinking water were found to have delayed onset of B-ALL at 11 weeks of age and changes in gene expression relating to B cell development, calcium-apoptosis signaling, and pathways in cancer, although no differences in lifespan were observed. Tumours from treated …

Effects Of Annexin A5 On Endothelial Inflammation Induced By Lipopolysaccharide-Activated Platelets And Extracellular Vesicles, Brent Jeffrey Tschirhart

Electronic Thesis and Dissertation Repository

Sepsis is a dysregulated immune response to infection and the leading cause of mortality globally, accounting for 11 million deaths in 2017. To date, no therapeutics are available to treat the underlying septic response. Previous research from our laboratory has shown that annexin A5 (Anx5) treatment increased survival by 40% in mice with endotoxemia, a model of sepsis. During sepsis, activated platelets release membrane fragments called extracellular vesicles (EVs) with externalization of phosphatidylserine to which annexin A5 binds with a high affinity. We hypothesized that annexin A5 will block the pro-inflammatory response induced by activated platelets and EVs in vascular …

Testing A Novel Peptide-Infused Cream For Treatment Of Rheumatoid Arthritis In Dr4tg Mice, Alisha J. Moynahan

Electronic Thesis and Dissertation Repository

Rheumatoid arthritis (RA) is a chronic inflammatory autoimmune disease that causes joint pain and damage. Studies have shown that inducing immune tolerance towards RA-specific proteins/peptides in RA mouse models can reduce arthritis severity and pro-inflammatory responses. The objectives of this study were to determine if a novel peptide cream treatment could modify RA-specific immune responses and reduce joint swelling in a humanized mouse model expressing the HLA-DRB1*0401 allele (known as DR4tg mice), the strongest genetic risk-factor for RA. Hyaluronan-Phosphatidylethanolamine cream infused with synthetic peptides HomoCitJED and CitJED was applied to HomoCitJED-immunized DR4tg mice before or after arthritis induction. Knee joint …

The Role Of Regulator Of G Protein Signaling 2 In Inflammatory Cytokine Release In Endotoxemia In Mice, Xin Tong Ma

Electronic Thesis and Dissertation Repository

In sepsis, lipopolysaccharide (LPS) activates toll-like receptor 4 to stimulate the release of inflammatory cytokines (e.g. tumor necrosis factor-alpha, TNF-α), leading to cardiac dysfunction. Regulator of G protein signaling 2 (RGS2) limits G protein-coupled receptor signaling by increasing the rate of G protein deactivation or inhibiting G protein-effector interactions. We hypothesized that RGS2 deficiency would enhance proinflammatory responses in endotoxemia. Adult wild-type and RGS2^-/- C57BL/6 mice and neonatal cardiomyocytes were treated with LPS and assessed for inflammatory responses and cardiac function. Myocardial TNF-α expression was higher in RGS2^-/- mice during endotoxemia. Additionally, cardiac function was impaired in …

Characterizing T Cell Phenotype In Patients With Hypersensitivity Reactions To Sulfamethoxazole And Beta-Lactam Antibiotics, Christine Caron

Electronic Thesis and Dissertation Repository

Delayed drug hypersensitivity reactions (DHRs) are idiosyncratic, T-cell mediated, and can present days after exposure to the culprit drug, resulting in varying degrees of skin rashes. We hypothesize that differences in activated peripheral T cell subsets and types of mediators released produce different clinical phenotypes of drug hypersensitivity reactions to sulphnamides and beta-lactam antibiotics.

We recruited participants with previous DHRs to sulfamethoxazole or beta-lactams . Peripheral blood mononuclear cells were isolated from participants. T-cell subset proliferation and activation was assessed by T-cell specific surface markers using 3H- thymidine incorporation and flow cytometry, and secreted cytokines were measured using bead-based detection. …

Development And Evaluation Of A Heterogenous Virus-Like Particle (Vlp) Formulation To Achieve Hiv-1 Latency Reversal And Cure., Joshua P. Pankrac

Electronic Thesis and Dissertation Repository

HIV-1 is the etiological agent behind acquired immune deficiency syndrome (AIDS) – a chronic, life-threatening condition that compromises host immune function. After nearly four decades and despite ongoing global efforts, HIV-1 persists in nearly 38 million individuals worldwide. Of this population, only 60% have access to life-saving combination antiretroviral therapy (cART), clearly emphasizing the need to realize a cure. Unfortunately, the establishment of replication-competent provirus in resting CD4+ T lymphocytes represents a significant barrier to HIV-1 curative research. The viral reservoir is highly stable and has a half-life of ~44 months. Therefore, it is unlikely that infection will naturally exhaust …

Utilizing Fiv (Feline Immunodeficiency Virus) To Develop A Novel Animal Model To Study Hiv (Human Immunodeficiency Virus), Ankita Suryakant Kambli

Electronic Thesis and Dissertation Repository

This project sought to perform the in vitro work needed to accomplish the long-term vision of harnessing the similarities between HIV (Human Immunodeficiency Virus) and FIV (Feline Immunodeficiency Virus) to develop an animal model whereby cats can be used to study HIV pathogenesis and therapeutics. We transfected CRFK (Crandell Rees Feline Kidney) fibroblasts with plasmids that could express human or feline CD4, CCR5, or both, and determined receptor surface expression through flow cytometry. We discovered that HIV envelope expressed on 293T can fuse with huCD4/huCCR5 on CRFK. These cat cell lines were also capable of supporting HIV infection. Additionally, we …

Determining The Relative Transmission Fitness Of Hiv-1 Subtypes A, B, C, And D, Spencer Yeung

Electronic Thesis and Dissertation Repository

There is in vivo evidence that suggests the genetic diversity of HIV-1 subtypes influence heterosexual transmission efficiency. To recapitulate sexual transmission in vitro, blocks of genital tissue were exposed to mixtures of genetically different subtype viruses. Migrating immune cells were collected and co-cultured with a CD4+ T-cell line permissive to HIV infection (PM1) to measure dendritic cell virus transfer; HIV-exposed tissues were cultured separately. Next generation sequencing (NGS) of HIV-1 DNA was used to quantify relative infection rates of the various challenge viruses, and to assess fitness differences in infection of the tissue vs. migratory/T cell co-cultures. Our results …

Hiv-1 Group M Subtype Fitness, Disease Progression, And Entry Efficiency, Colin M. Venner

Electronic Thesis and Dissertation Repository

Human immunodeficiency virus type 1 (HIV-1) emerged in the human population shortly after the turn of the 19th century. Distribution of HIV-1 across the globe over the past 30–35 years can be traced to founder events with primordial HIV strains from sub-Saharan Africa. Even considering the burden of HIV in Africa, our knowledge of HIV-1 disease is still largely limited to subtype B HIV-1, a strain responsible for 3 million infections in North America and Europe as compared to the 33 million that are infected with HIV-1 subtypes A, C, D, and circulating and unique recombinant forms.

This dissertation analyzes …

Effects Of G Protein Signalling Modulator 3 On Cellular Signalling, Aneta A. Surmanski

Electronic Thesis and Dissertation Repository

G protein coupled receptors (GPCRs) promote G protein heterotrimer (Gα·GDP/Gbg) activation.GPCRsignalling is limited via G protein GTPase activity and b-arrestin-receptor interactions. G Protein Signalling Modulators (GPSMs) are proteins that may influence receptor signalling through G protein activity. GPSM3 modulates their activity by binding to Ga_i-GDP, limiting nucleotide exchange and preventing its re-association to Gbg. The impact of GPSM3 on signalling is unknown.We hypothesize that GPSM3 will decrease Ga_i-dependent signalling while promoting Gbg-dependent signalling in G_i-coupled GPCRs.

GPSM3 significantly inhibited b-arrestin recruitment to α_2A-adrenergic and m-opioid receptors via a Gbg-dependent mechanism, …

Cd4+ T-Cell Mediated Microvascular Endothelial Cell Death And Chronic Cardiac Allograft Rejection Involves Necroptosis, Cecilia Yt Kwok

Electronic Thesis and Dissertation Repository

Heart transplantation is the only viable option for patients with end-stage heart failure. Despite advances in immunosuppressive therapies, the rate of transplanted graft loss remains substantial. Graft loss is primarily due to tissue damage mediated by immune responses. Cell death and organ rejection can occur as an active molecular process through apoptotic and necrotic pathways. We now recognize that cell death may also ensue through a newly described form of programmed necrotic cell death, termed necroptosis that involves receptor-interacting protein kinase (RIPK) 1/3. In this study, I aim to establish the role of RIPK3 in T cell-mediated chronic cardiac allograft …

Granulocyte Colony-Stimulating Factor: Its Role In Gut-Homing Macrophage Generation And Colitis, And Production By Probiotics, Shahab Meshkibaf

Electronic Thesis and Dissertation Repository

The pleiotropic cytokine granulocyte-colony stimulatory factor (G-CSF) is mainly required for the generation of neutrophils, but its role in macrophage generation has also been reported. In addition, G-CSF is effective for the down-regulation of inflammatory cytokines and ameliorating gut disorders, such as colitis. However, the G-CSF function in macrophage generation and gut immunity remains unclear. The first focus of this thesis was to assess the role of G-CSF in macrophage generation and its contribution to gut immunity. G-CSF was found to promote the generation of Gr-1high/F4/80+ macrophages in macrophage (M)-CSF-treated bone marrow cells, most likely through suppressing cell death. Gr-1high …

Methotrexate And Cardiovascular Events, Alpesh Shah

Electronic Thesis and Dissertation Repository

We conducted a systematic review and meta-analysis to assess the association of MTX with cardiovascular morbidity, cardiovascular mortality and all-cause mortality in patients with autoimmune disease. Our primary outcome was incident cardiovascular events. After screening 13,479 citations, we identified a total of 30 eligible studies. We synthesized adjusted risk estimates using a random effects model. MTX was significantly associated with a 25% reduction in cardiovascular events (pooled RR: 0.75, 95% CI: 0.65, 0.86, I²: 11%), a 55% reduction in cardiovascular mortality (0.45, 95% CI: 0.26, 0.80, I²: 33%) and a 40% reduction in all-cause mortality (0.60, …

Cell Death Regulates Injury And Inflammation During Renal Allograft Transplantation, Arthur Lau

Electronic Thesis and Dissertation Repository

Renal transplantation invariably results in tissue injury resulting from ischemia reperfusion injury (IRI), inflammation, drug toxicity, and rejection. Tubular epithelial cells (TEC) comprise the majority of renal parenchyma and are susceptible to cell death and injury during diverse forms of inflammation, which has direct and indirect effects on long term allograft function. Renal TEC have the unique ability to attenuate inflammation and alloimmune injury through the expression of various mediators of cell death and inflammatory molecules. Inhibition of cell death pathways in renal allografts may influence outcomes of alloimmune responses and graft survival. In this body of investigation, alteration of …

Analysis Of Invariant Natural Killer T Cells In Intra-Abdominal Sepsis, Ram Venkatesh Anantha

Electronic Thesis and Dissertation Repository

Sepsis is characterized by a severe systemic inflammatory response to infection that is associated with high morbidity and mortality despite optimal care. Invariant natural killer T (iNKT) cells are potent regulatory lymphocytes that can produce pro- and/or anti-inflammatory cytokines, thus shaping the course and nature of immune responses; however, little is known about their role in sepsis. We demonstrate here that patients with sepsis/severe sepsis have significantly elevated proportions of circulating iNKT cells in their peripheral blood, as compared to non-septic patients. We therefore investigated iNKT cells in mice with intra-abdominal sepsis (IAS). Our data show that iNKT cells are …

The Role Of Il-22 Produced By Th17 Cells In Type 1 Diabetes, Stacey M. Bellemore

Electronic Thesis and Dissertation Repository

Interleukin-22 (IL-22) is produced by T helper 17 (Th17) cells. Th17 cells have been shown to be pathogenic in autoimmune diseases, however their role in type 1 diabetes (T1D) remains controversial. We have shown that Th17-differentiation of naïve T cells can be driven by IL-23 + IL-6 to produce large amounts of IL-22 and induce T1D. Conversely, polarizing T cells using TGF-β + IL-6 led to nonpathogenic Th17 cells that produced lower IL-22 levels. We have shown that neutralizing IFN-γ during polarization leads to a drastic increase in IL-22. We have also found IL-22-producing cells in the pancreas of diabetic …

Role Of Inos In Septic Pulmonary Microvascular Endothelial Cell Activation, Zahra Asad

Electronic Thesis and Dissertation Repository

Abstract

Background. Neutrophils and nitric oxide (NO) derived from inducible NO synthase (iNOS) contributes importantly to the pathophysiology of acute lung injury (ALI) and pulmonary microvascular endothelial cell (PMVEC) injury. However, the mechanism of neutrophil and neutrophil iNOS dependent PMVEC injury has not been addressed. In our studies, we assessed PMVEC activation under septic conditions, and defined the role of PMVEC vs. bone-marrow polymorphonuclear leukocytes (PMN) iNOS in this septic PMVEC activation.

Methods and Results. We isolated PMVEC from iNOS+/+ and iNOS-/- mice lungs magnetically by microbeads attached to anti-PECAM antibodies, sorted by flow cytometry (FACS) by DiI-acetylated low density …

Immune Responses To Homocitrullinated Protein/Peptide In Rheumatoid Arthritis, Mathias J. Scinocca

Electronic Thesis and Dissertation Repository

Rheumatoid Arthritis (RA) is an inflammatory autoimmune disease which causes joint destruction. RA pathogenesis involves citrullinated peptides binding to the shared epitope (SE) during autoantigen presentation, and subsequent Anti-Citrulline Antibody (ACA) production. Their target, citrulline, is very similar to homocitrulline.

The main objective of this study was to investigate anti-homocitrulline immune responses in RA. Specifically, it investigated if: i) Anti-Homocitrulline Antibodies (AHA) were RA specific by screening patients with various inflammatory rheumatic diseases and healthy individuals. ii) ACA also bound homocitrulline by affinity purification and characterization. iii) anti-homocitrulline immune responses involved the SE by computer modelling and immunization of mice. …

Restriction Of Hiv-1 Replication By Unique Trim22 Isoforms., Clayton Hattlmann

Electronic Thesis and Dissertation Repository

Understanding how the immune system reacts to HIV infection and why normal antiviral defenses are insufficient to fight infection is a key step towards creating better therapies. Several interferon-induced proteins, such as the tripartite motif protein TRIM22, are capable of restricting HIV-1 replication; however single nucleotide polymorphisms (SNPs) can dramatically impact the actions of these proteins. While the trim22 gene contains numerous SNPs, no study has addressed how these may affect TRIM22 functions. Here we provide the first direct comparison of two TRIM22 unique isoforms. Through confocal microscopy we observed these isoforms exhibit different patterns of localization. In vitro studies …